DNA双链断裂作为α粒子发射体Ac-225和电子发射体Lu-177用于生长抑素受体靶向放射治疗疗效的预测指标。
DNA double strand breaks as predictor of efficacy of the alpha-particle emitter Ac-225 and the electron emitter Lu-177 for somatostatin receptor targeted radiotherapy.
作者信息
Graf Franziska, Fahrer Jörg, Maus Stephan, Morgenstern Alfred, Bruchertseifer Frank, Venkatachalam Senthil, Fottner Christian, Weber Matthias M, Huelsenbeck Johannes, Schreckenberger Mathias, Kaina Bernd, Miederer Matthias
机构信息
University Medical Centre, Department of Nuclear Medicine, Mainz, Germany.
University Medical Centre, Institute of Toxicology, Mainz, Germany.
出版信息
PLoS One. 2014 Feb 7;9(2):e88239. doi: 10.1371/journal.pone.0088239. eCollection 2014.
RATIONALE
Key biologic effects of the alpha-particle emitter Actinium-225 in comparison to the beta-particle emitter Lutetium-177 labeled somatostatin-analogue DOTATOC in vitro and in vivo were studied to evaluate the significance of γH2AX-foci formation.
METHODS
To determine the relative biological effectiveness (RBE) between the two isotopes (as - biological consequence of different ionisation-densities along a particle-track), somatostatin expressing AR42J cells were incubated with Ac-225-DOTATOC and Lu-177-DOTATOC up to 48 h and viability was analyzed using the MTT assay. DNA double strand breaks (DSB) were quantified by immunofluorescence staining of γH2AX-foci. Cell cycle was analyzed by flow cytometry. In vivo uptake of both radiolabeled somatostatin-analogues into subcutaneously growing AR42J tumors and the number of cells displaying γH2AX-foci were measured. Therapeutic efficacy was assayed by monitoring tumor growth after treatment with activities estimated from in vitro cytotoxicity.
RESULTS
Ac-225-DOTATOC resulted in ED50 values of 14 kBq/ml after 48 h, whereas Lu-177-DOTATOC displayed ED50 values of 10 MBq/ml. The number of DSB grew with increasing concentration of Ac-225-DOTATOC and similarly with Lu-177-DOTATOC when applying a factor of 700-fold higher activity compared to Ac-225. Already 24 h after incubation with 2.5-10 kBq/ml, Ac-225-DOTATOC cell-cycle studies showed up to a 60% increase in the percentage of tumor cells in G2/M phase. After 72 h an apoptotic subG1 peak was also detectable. Tumor uptake for both radio peptides at 48 h was identical (7.5%ID/g), though the overall number of cells with γH2AX-foci was higher in tumors treated with 48 kBq Ac-225-DOTATOC compared to tumors treated with 30 MBq Lu-177-DOTATOC (35% vs. 21%). Tumors with a volume of 0.34 ml reached delayed exponential tumor growth after 25 days (44 kBq Ac-225-DOTATOC) and after 21 days (34 MBq Lu-177-DOTATOC).
CONCLUSION
γH2AX-foci formation, triggered by beta- and alpha-irradiation, is an early key parameter in predicting response to internal radiotherapy.
原理
研究了α粒子发射体锕-225与β粒子发射体镥-177标记的生长抑素类似物多他曲妥（DOTATOC）在体外和体内的关键生物学效应，以评估γH2AX焦点形成的意义。
方法
为了确定两种同位素之间的相对生物学效应（RBE）（作为沿粒子轨迹不同电离密度的生物学后果），将表达生长抑素的AR42J细胞与Ac-225-DOTATOC和Lu-177-DOTATOC孵育长达48小时，并使用MTT法分析细胞活力。通过γH2AX焦点的免疫荧光染色对DNA双链断裂（DSB）进行定量。通过流式细胞术分析细胞周期。测量了两种放射性标记的生长抑素类似物在皮下生长的AR42J肿瘤中的体内摄取情况以及显示γH2AX焦点的细胞数量。通过监测根据体外细胞毒性估计的活性进行治疗后肿瘤的生长情况来测定治疗效果。
结果
48小时后，Ac-225-DOTATOC的半数有效剂量（ED50）值为14 kBq/ml，而Lu-177-DOTATOC的ED50值为10 MBq/ml。DSB的数量随着Ac-225-DOTATOC浓度的增加而增加，当应用比Ac-225高700倍的活性时，Lu-177-DOTATOC的情况类似。与2.5 - 10 kBq/ml孵育24小时后，Ac-225-DOTATOC的细胞周期研究显示G2/M期肿瘤细胞百分比增加高达60%。72小时后也可检测到凋亡的亚G1峰。48小时时两种放射性肽的肿瘤摄取相同（7.5%ID/g），尽管与用30 MBq Lu-177-DOTATOC治疗的肿瘤相比，用48 kBq Ac-225-DOTATOC治疗的肿瘤中显示γH2AX焦点的细胞总数更高（35%对21%）。体积为0.34 ml的肿瘤在25天（44 kBq Ac-225-DOTATOC）和21天（34 MBq Lu-177-DOTATOC）后达到延迟的指数生长。
结论
由β和α辐射引发的γH2AX焦点形成是预测内放疗反应的早期关键参数。
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